The ever-increasing number of bone defects arising from clinical disorders and trauma injuries constitutes a significant healthcare concern and is a costly source of annual expenditure. The finite supply of autologous bone grafts, incompatibility of allogeneically sourced bone, and poor integration within large defects provide the impetus for developing viable bone repair alternatives. To that end, tissue engineering strategies integrating cells, biomaterials, and/or bioactive factors have demonstrated tremendous progress toward regenerating bone. While bone morphogenetic protein-2 (BMP-2)'s osteoinductivity is frequently capitalized upon, its potency often compromises the integrity of regenerated bone. Despite advances in biomaterial- and gene- based controlled release approaches, precise BMP-2 delivery over a desired duration remains a critical challenge. Translational research strategies that improve regulation of BMP-2 expression levels could therefore spur the development of safer and more efficacious treatments for bone defects unable to heal through natural processes. Herein we propose the design of an innovative red/far red (R/FR) light- controlled BMP-2 release platform to promote osteogenesis in vitro and in vivo. Optical regulation is advantageous over currently existing platforms due to its non-invasiveness, analog tunability, and robust spatiotemporal resolution. Our technology will leverage recently developed optogenetic tools merging light- responsive protein domains Phytochrome B (PhyB) and Phytochrome Interacting Factor-6 (PIF6) from Arabidopsis thaliana with engineered gene regulatory elements. When illuminated by red (660nm) light, the PhyB/PIF6 system activates a downstream promoter, which can then be instantaneously turned off by far red (740nm) light. To our knowledge, no prior studies have investigated the use of these modules for tissue engineering applications. The long-term goal of this proposal entails the development of a cell-based controlled release platform in which the concentration and timeframe of a delivered therapeutic factor can be programmed via specific wavelengths of light. The objective is to derive a pool of stable rat mesenchymal stem cells (MSCs) harboring a R/FR light gene regulatory circuit for BMP-2 release. The central hypothesis is that R/FR light ratios will permit analog tuning of BMP-2 levels, and that the resulting optical precision will improve bone formation. In Specific Aim 1, we will engineer and characterize a BMP-2 expressing R/FR light-controlled gene regulatory circuit stably integrated in rat MSCs. In Specific Aim 2, we will evaluate the osteogenic potential of these genetically modified cells in vitro and in vivo using R/FR light ratios selected from Specific Aim 1 and different biomaterial carriers. If successful, the proposed research will yield a clinically relevant cell- based therapeutic factor release platform for bone regeneration, as well as a design framework translatable to other tissue engineering applications.